被孢霉W_（A－90）发酵生产γ－亚麻酸的研究

本文研究了由被抱霉变株ＭＡ－９０生产了γ－亚麻酸的发酵条件．确定了最适碳源、复源及Ｃ／Ｎ．初步建立了生产γ－亚麻酸的工艺条件。葡萄糖、蔗糖及天冬酰胺和尿素为最适碳、氮源。在Ｃ／Ｎ为２０／１,葡萄糖浓度为８０ｇ／Ｌ的条件下．油脂产量和油脂中γ－亚麻酸的含量分别为８．２ｇ／Ｌ和１４．１３％,γ－亚麻酸产量及生物量分别为１．１５７ｇ／Ｌ和３１．２ｇ／Ｌ。后期适当降低培养温度和良好的通气条件均有利于γ－亚麻酸的积累。     

Specificity studies of 3-mercaptopyruvate sulfurtransferase

3-Mercaptopyruvate sulfurtransferase (E.C. 2.8.1.2; MST) is an enzyme believed to function in the endogenous cyanide (CN) detoxification system because it is capable of transferring sulfur from 3-mercaptopyruvate (3-MP) to CN, forming the less toxic thiocyanate (SCN). To date, 3-MP is the only known sulfur-donor substrate for MST. In an effort to increase the understanding of what chemical properties of 3-MP affect its utilization as a substrate, in vitro enzyme kinetic studies of MST were conducted using two mercaptic acids that are structurally related to 3-MP. Neither of these compounds was able to serve as a sulfur-donor substrate for MST. Inhibitor studies determined that 3-mercaptopropionic acid did not affect the K_m of MST for 3-MP but did decrease V_max and, thus, was determined to be a noncompetitive inhibitor. Alternatively, 2-mercaptopropionic acid 2-MPA decreased K_m and V_max and was determined to be an uncompetitive inhibitor of MST with respect to 3-MP. These data indicate that the α-keto group of 3-MP is necessary for its utilization as a substrate, and the inhibitor studies suggest that the position of the sulfur may also affect the binding of these compounds to the enzyme. These observations increase the understanding of what factors can affect the utilization of a compound as a sulfur-donor substrate for MST and may aid in the development of alternative sulfur-donor substrates for MST. © 1996 John Wiley & Sons, Inc.     

The adaptive acid tolerance response in root nodule bacteria and Escherichia coli

Root nodule bacteria and Escherichia coli show an adaptive acid tolerance response when grown under mildly acidic conditions. This is defined in terms of the rate of cell death upon exposure to acid shock at pH 3.0 and expressed in terms of a decimal reduction time, D. The D values varied with the strain and the pH of the culture medium. Early exponential phase cells of three strains of Rhizobium leguminosarum (WU95, 3001 and WSM710) had D values of 1, 6 and 5 min respectively when grown at pH 7.0; and D values of 5, 20 and 12 min respectively when grown at pH 5.0. Exponential phase cells of Rhizobium tropici UMR1899, Bradyrhizobium japonicum USDA110 and peanut Bradyrhizobium sp. NC92 were more tolerant with D values of 31, 35 and 42 min when grown at pH 7.0; and 56, 86 and 68 min when grown at pH 5.0. Cells of E. coli UB1301 in early exponential phase at pH 7.0 had a D value of 16 min, whereas at pH 5.0 it was 76 min. Stationary phase cells of R. leguminosarum and E. coli were more tolerant (D values usually 2 to 5-fold higher) than those in exponential phase. Cells of R. leguminosarum bv. trifolii 3001 or E. coli UB1301 transferred from cultures at pH. 7.0 to medium at pH 5.0 grew immediately and induced the acid tolerance response within one generation. This was prevented by the addition of chloramphenicol. Acidadapted cells of Rhizobium leguminosarum bv. trifolii WU95 and 3001; or E. coli UB1301, M3503 and M3504 were as sensitive to UV light as those grown at neutral pH.     

Carbon allocation in developing spruce needles. Enzymes and intermediates of sucrose metabolism

In lyophilized needles of Norway spruce ( Picea abies [L.] Karsten) and starting from bud break, we determined enzyme activities (sucrose phosphate synthase [SPS; EC 2.4,1.14]. sucrose synthase [SS; EC 2.4,1.13]. acid invertase [AI; EC 3.2,1.26]) and intermediates (starch, sucrose, glucose, fructose; fructose 6-phosphate, fructose 2.6-bisphosphate [F26BP]) of carbohydrate metabolism together with needle weight, shoot length, chlorophyll and protein. For up to 110 days after bud break, samples were taken twice a week from about 25-year-old trees under field conditions. At least three periods can be distinguished during needle maturation. During the first period (up to 45 days after bud break) Al showed the highest extractable activity. This coincided with very high levels of F26BP (up to 11 pmol [mg dry weight]⁻¹) and a transient increase of starch in parallel to a decrease of sucrose. The interval between 45 and 70 days after bud break was characterized by high SS activity (ratio of fructose/glucose >1), much decreased levels of F26BP (down to below 1 pmol [mg dry weight]⁻¹), and a pronounced increase in the dry weight/fresh weight ratio. In parallel, starch declined and soluble carbohydrates increased. Finally, needle maturation was characterized by decreasing SS and continuously increasing SPS activities, so that the ratio of SPS/SS increased more than 6-fold. AI. however, did not decline with maturation. Changes in pool sizes of metabolites and enzyme activities (AI. SPS) are consistent with current concepts on sink/source transition. SS is obviously important with regard to the synthesis of structural polysaccharides.     

The role of acid phosphatases in plant phosphorus metabolism

Hydrolysis of phosphate esters is a critical process in the energy metabolism and metabolic regulation of plant cells. This review summarizes the characteristics and putative roles of plant acid phosphatase (APase). Although immunologically closely related, plant APases display remarkable heterogeneity with regards to their kinetic and molecular properties, and subcellular location. The secreted APases of roots and cell cultures are relatively non-specific enzymes that appear to be important in the hydrolysis and mobilization of P_i from extracellular phosphomonoesters for plant nutrition. Intracellular APases are undoubtedly involved in the routine utilization of P_i reserves or other P_i-containing compounds. A special class of intracellular APase exists that demonstrate a clear-cut (but generally nonabsolute) substrate selectivity. These APases are hypothesized to have distinct metabolic functions and include: phytase, phosphoglycolate phosphatase, 3-phosphoglycerate phosphatase, phosphoenolpyruvate phosphatase, and phosphotyrosyl-protein phosphatase. APase expression is regulated by a variety of developmental and environmental factors. P_i starvation induces de novo synthesis of extra- and intracellular APases in cell cultures as well as in whole plants. Recommendations are made to achieve uniformity in the analyses of the different APase isoforms normally encountered within and between different plant tissues.     

S-PI对香菇酸性蛋白酶的抑制机理及其促进出菇的生化解释

S-PI对纯化的香菇酸性蛋白酶具有强烈的抑制作用,10μmol／L的S-PI可使酶活降低77％,这种抑制属于可逆竞争抛物线型抑制。经25μmol／L酸性蛋白酶水解6小时后,香菇胞外酸性磷酸酶的活性从24．6单位降低到20．8单位,同时C1酶和Cx酶的活性分别比对照降低了76％和58％,这表明香菇酸性蛋白酶不仅直接左右着香菇发育的氮代谢,同时也间接地影响着碳代谢及能量转换。     

金属离子螯合剂对橙汁中L—抗坏血酸的保护及其机理探讨

本文研究了在加热过程中金属离子螯合剂植酸、聚磷酸钠和EDTANa_2对橙汁和橙汁模拟体系中L—抗坏血酸的稳定作用,并采用红外光谱和紫外差光谱对EDTANa_2的保护机理作了初步探讨。结果表明:植酸和聚磷酸钠均不能降低Cu~(2+)对橙汁模拟体系中L—抗坏血酸氧化降解的催化作用,而EDTANa_2不仅能络合Cu~(2+),减少L—抗坏血酸的催化降解损失,而且红外光谱和紫外差光谱及溶剂微扰研究还揭示出EDTANa_2可与L—抗坏血酸形成氢键保护,这种氢键保护作用受到糖和柠檬酸的不利影响。     

急性肝损伤大鼠模型的血浆氨基酸代谢变化模式

以Ｄ－氨基半乳糖（Ｄ－Ｇａｌａｃｔｏｓａｍｉｎｅ,Ｄ－ＧａｌＮ）造成急性肝损伤（急性肝炎、急性肝坏死）大鼠模型后、对照观察了急性肝损伤大鼠血浆氨基酸的变化,建立了大鼠急性肝损伤时血浆氨基酸的变化模式并对其发生机理进行了探讨。大鼠血浆氨基酸的测定采用聚酰薄层荧光分析技术,其测定结果是：急性肝炎组,酪氨酸（Ｔｙｒ）、天冬氨酸（Ａｓｐ）、谷氨酰胺（Ｇｌｎ）和鸟氨酸（Ｏｒｎ）升高,精氨酸（Ａｒｇ）下降,其余氨基酸无显著变化。急性肝坏死组,除Ａｒｇ显著下降外,其余所有氨基酸都显著升高,而两组支链氨基酸（ＢＣＡＡ）／芳香族氨基酸（ＡＡＡ）克分子比值均显著下降。     

Purification and partial characterization of an acid β-fructosidase from sweet-pepper (Capsicum annuum L.) fruit

The present study describes the biochemical characteristics of an acid -fructosidase (EC 3.2.1.26) purified from the fruit of sweet pepper (Capsicum annuum L.). The soluble form, which constitutes more than 95% of the total activity at pH 4.5, hydrolyzes sucrose, raffinose, and stachyose. Its pH and temperature optima are 4.5 and 55 °C, respectively. Metal cations such as Ag⁺ and Hg²⁺ strongly inhibit its activity, suggesting the presence of at least one sulfhydryl group at the catalytic site. After purification of the enzyme by means of ammonium sulfate fractionation, gel chromatography (diethyl-aminoethyl-Sephacel, hydroxylapatite, concanavalin A-Sepharose), and preparative gel electrophoresis, the purified enzyme was shown to be a 42 kDa glycoprotein interacting specifically with concanavalin A. After complete chemical deglycosylation with trifluoromethanesulfonic acid, the molecular weight of the constitutive polypeptide was estimated to be 39 kDa. The enzyme glycans were characterized using both affino- and immunodetection. The enzyme has at least two N-linked oligosaccharide sidechains, one of the high-mannose type, and the other of the complex type. The high-mannose glycan has a low molecular weight (1 kDa), and is responsible for the interaction between the enzyme and concanavalin A. The complex-type glycan has an estimated molecular weight of 2 kDa. It contains one 1 2-linked xylose residue, probably one fucose residue 1 3-linked to the chitobiose unit, and no terminal galactose residue. The two glycans, associated to the 39 kDa polypeptide, constitute the acid -fructosidase of the sweet-pepper fruit.Abbreviations F -fructosidase - ConA concanavalin A - DEAE diethylaminoethyl - DTNB dithionitrobenzoic acid - endo F endo--N-acetylglucosamidase F - endo H endo--N-acetylglucosamidase H - NEM N-ethylmaleimide - PCMB parachloromercurobenzoate - PNGase glycopeptide-N-glycosidase - TFMS trifluoromethane sulfonic acid This work was partly supported by a grant from the Commission Permanente de Coopération Franco-Québécoise to L. Faye, and S. Yelle. D. Michaud was a recipient of a graduate scholarship from the Natural Science and Engineering Research Council of Canada.